The dramatic increase in healthcare costs has become a significant burden to the world, with many patients being denied medications because of their high prices. The biosynthesis of chiral drugs and drug intermediates offers an environmentally friendly approach to addressing such problems, for example, by providing cost effective methodologies for the production of therapeutic agents as well as the intermediates and/or precursors used to make such agents.
L-homoalanine is a nonnatural amino acid that is a key chiral intermediate for the synthesis of several important drugs (FIG. 1A). For example, it can be converted to S-2-aminobutyramide, which is the immediate precursor of the antiepileptic drugs levetiracetam and brivaracetam. L-homoalanine can also be converted to S-2-aminobutanol, a chemical intermediate in methods for synthesizing the antituberculosis compound ethambutol. Methods for synthesizing therapeutic compounds such as levetiracetam and ethambutol must overcome a number of technical challenges. For example, the optical purity of these drugs is critical for therapeutic safety and efficacy. The R-enantiomer of levetiracetam has no antiepileptic activity (see, e.g. Shorvon et al., (2002) Journal of Neurology, Neurosurgery, and Psychiatry 72(4):426-429) and (R,R)-form of ethambutol can cause blindness (see, e.g. Breuer M, et al. (2004) Angewandte Chemie International Edition 43(7):788-824).
Even though ethambutol and levetiracetam are now generic drugs, in many countries the cost of just one month's supply exceeds the entire annual per capita health expenditure (see, e.g. Moore-Gillon J (2001) Ann NY Acad Sci 953:233-240). The prohibitive drug price has created global healthcare problems. For example, while epilepsy affects over 50 million people worldwide, most of the patients cannot afford the levetiracetam treatment, and must use cheaper but much less effective alternatives such as phenobarbital (see, e.g. Scott et al. (2001) B World Health Organ 79:344-351). One approach to reducing drug costs in order to make them more widely available involves cost-effective approaches to L-homoalanine synthesis (e.g. by reducing the manufacturing cost of compounds such as levetiracetam).
Most of the natural L-amino acids can now be produced from glucose by microbial fermentation (see, e.g. Ikeda M (2002) Adv Biochem Eng Biot 79:1-35). Notably, L-glutamate, L-lysine, and L-threonine are produced more than 2 million tons annually (see, e.g. Leuchtenberger et al. (2005) Appl Microbiol Biotechnol 69(1):1-8). In contrast to methods for making natural L-amino acids, methods for the commercial-scale preparation of nonnatural amino acids are typically complex as well as environmentally unfriendly. In one prior art approach, chemically synthesized 2-ketoacids are asymmetrically converted to optically pure nonnatural amino acids by transaminases or dehydrognenases (see, e.g. Leuchtenberger et al. (2005) Appl Microbiol Biotechnol 69(1):1-8; Taylor et al. (1998) Trends Biotechnol 16(10):412-418). Another approach uses enzymes such as acylases or amidases to resolve racemic mixtures of nonnatural amino acids (see, e.g. Leuchtenberger et al. (2005) Appl Microbiol Biotechnol 69(1):1-8).
Due to, for example, their usefulness in a making a variety of valuable therapeutic compounds, there is a need in the art for methods and materials that facilitate the cost effective and environmentally friendly biosynthesis of nonnatural amino acids such as L-homoalanine. Unlike natural amino acids however, the total biosynthesis of nonnatural amino acids from simple sugars involves significant technical challenges. For example, in one environmentally friendly and cost effective approach, metabolic pathways in an organism are altered in order to expand the biosynthetic capabilities of that organism (see, e.g. Zhang et al. (2008) Proc Natl Acad Sci USA 105(52):20653-20658). In such approaches, the altered metabolic pathways then facilitate or direct the production of a target compound such as a nonnatural amino acid (see, e.g. Causey et al. (2003) Proc Natl Acad Sci USA 100(3):825-832). Unfortunately, however, the results of any manipulation designed to alter an organism's metabolic pathways can be unpredictable and such efforts typically require extensive protein evolution (see, e.g. Arnold F H (2001) Nature 409(6817):253-257).